Metabolism and renal clearance of In-111-labeled DOTA-conjugated antibody fragments

Radiometal-labeled antibody fragments are promising reagents for radioimmun otherapy due to their high tumor uptake and rapid pharmacokinetics, but the ir therapeutic potentials are limited by high uptake and retention in the k idney. Identification of metabolic products is a first step in designing ra tionale approaches to lower kidney uptake. Previous studies in rats have sh own that In-111-labeled DTPA-conjugated antibody fragments (via lysine resi dues) were degraded to an DTPA-is an element of -amino-lysine derivative an d retained in the lysosomal compartments of the liver and kidney [Rogers et al. (1995) Cancer Res. 55, 5714s-5720s]. To determine the metabolic profil e of another widely used metal-chelate, [In-111]DOTA conjugated to lysines in antibody fragments via active ester chemistry, we analyzed kidney homoge nates from nude mice injected with an [In-111]DOTA-Fab generated enzymatica lly from the anti-lymphoma intact antibody Rituxan. The major kidney metabo lite was identified as [In-111]DOTA-is an element of -amino-lysine by compa rison to an authentic synthetic standard. This end product was also identif ied in the urine, along with relatively small amounts of [In-111]DOTA-Fab. Since injection of [In-111] DOTA-is an element of -amino-lysine into nude m ice resulted in rapid clearance into the urine without kidney retention, it is likely that the renal retention observed was due to kidney uptake of [I n-111]DOTA-Fab, followed by lysosomal degradation to [In-111]DOTA-is an ele ment of -amino-lysine, which is only slowly cleared from this compartment. This observation is supported by autoradiographs of the kidney showing rapi d localization of radioactivity into the distal regions of the kidney corte x. To extend this analysis to clinical trials, we have also analyzed urine taken from a patient injected with the intact antibody [In-111] DOTA-cT84.6 6. In that example, we found that the major radioactive species was also [I n-111] DOTA-is an element of -amino-lysine.